The project focuses on the elucidation of molecular mechanisms by which the immunologically important NF-kappaB transcription factors can be activated. Identification of the molecular components of signaling pathways will also reveal potential targets for therapeutic intervention to block activation of NF-kappaB. Inflammatory diseases are often driven by the undesirable activation of NF-kappaB. In addition, this factor figures prominently in the expression of the human immunodeficiency virus, and other clinically relevant viruses. The NF-kappaB transcription factors are normally retained in the cytoplasm by association with the inhibitory IkappaB proteins. Signals directly or indirectly related to pathogens or stress lead to phosphorylation and then proteolytic degradation of the inhibitor, thereby releasing the NF-kappaB factors to translocate to the nucleus and carry out their functions. Additional regulation is imposed via direct or indirect effects of signals on the transactivation activity of these factors once they have entered the nucleus. The present project is concerned with the delineation of several signaling paths that lead to phosphorylation and subsequent ubiquitin-dependent proteasomal degradation of the IkappaB inhibitors as well as those paths that stimulate transactivation. We have cloned CIKS, which is associated with the NEMO, the regulatory subunit of the core IkappaB kinase (IIK) complex. In addition to NEMO, the IKK core complex also contains the IkappaB kinases alpha and beta. Transfected CIKS not only activates NF-kappaB, but also the Jun Kinase. We have discovered that the IKK complex can be associated not only with CIKS, but also with two additional IKK-related kinases and with Tak1, a kinase implicated in IL-1 signal transduction. We have identified an adapter protein that may link the IKK-related kinases with the IKK core complex. These data provide evidence for a large complex containing several kinases and adapters that can surround the core IKK complex; these proteins are likely to be critical for funneling various different signals to the IKK kinases.